Surfactant proteins A and D (SP-A and SP-D) are important for innate defense against influenza A virus (lAV). We will examine the structural basis for neutralizing and opsonizing activities of these proteins. Minor changes in the carbohydrate recognition domain (CRD) near the lectin siteof SP-D markedly increase antiviral activity. Antiviral activity can be predicted by determining how amino acids on the CRD interact with mannose rich saccharide chains. Aim 1 examines binding of wild type and mutant neck+CRD (NCRD) trimer preparations of SP-D to whole virus, viral envelope preparations and purified glycans and correlates results with icrystallographic analysis and molecular modeling. Additional NCRDs with increased antiviral activity will be developed based on the results. Reverse genetics will be used to manipulate glycan attachments on the viral hemagglutinin to determine their role in neutralization by SP-D. We will also modify NCRDs based on properties of porcine SP-D which inhibits avian strains. Aim 2 examines how cooperative interactions between NCRD heads potentiates antiviral activity, using full length collectins with varied multimerization or collagen domain composition and NCRD preparations crosslinked through other means (i.e., antibodies). In addition to performing assays of viral binding and neutralization, aspects of antiviral activity that are most strongly affected by multimerization [viral aggregation and modulation of phagocyte interactions] will be examined. The role of the collectin collagen domain per se will also be tested. Methods of crosslinking NCRDs without use of the collagen domain can result in strong antiviral and opsonizing activity. This approach may be beneficial in vivo because receptors for the collagen domain of collectins are linked to pro inflammatory activities that could be harmful. We will finally use findings from aims 1 and 2 to construct collectins with optimal activity against human lAV strains and avian lAV strains. Both the NCRDs and full length recombinant collectins will be tested for activity in vivo in wild type and SP-D gene-deleted mice. Our dual goals are to clarify mechanisms of antiviral activity of collectins and to develop useful recombinant collectins for therapy of respiratory viruses. RELEVANCE (See instructions): This proposal examines certain natural proteins in the lung that protect against new influenza viruses. The proposal will clarify why some people are more subject to complications from influenza. In addition, new mutant proteins wiill be made that have stronger ability to inhibit human and avian types of influenza virus. These new proteins may be useful as treatments for influenza viruses in the future. PROJECT/PERFORIVIANCE SITE(S) (if additional space is needed, use Project/Performance